Keratin-containing inclusions affect cell morphology and distribution of cytosolic cellular components

Area of Science:

• Cell Biology
• Biochemistry
• Pathology

Background:

• Protein aggregates, including intermediate filament (IF) inclusions and Mallory bodies (MBs), are hallmarks of neurodegenerative and liver diseases.
• Intermediate filaments (IFs) anchor at desmosomes and hemidesmosomes, but their interactions with other intercellular junctions remain unclear.

Purpose of the Study:

• To investigate the impact of intermediate filament (IF) inclusions on junction-associated and cytosolic proteins within cultured cells.
• To understand how cytokeratin (CK) aggregation affects cellular morphology and protein localization.

Main Methods:

• Gene transfection of green fluorescent protein (GFP)-tagged cytokeratin 18 mutant (GFP-CK18 R89C) in cultured cells to induce CK aggregation and IF network loss.
• Immunofluorescence analysis to assess the colocalization of junction-associated proteins (zonula occludens-1, beta-catenin) and cytosolic proteins (14-3-3 zeta, glucose-6-phosphate dehydrogenase) with CK aggregates.
• Morphological assessment of transfected cells compared to controls.

Main Results:

• GFP-CK18 R89C transfection led to CK aggregation and disrupted IF networks.
• Zonula occludens-1, beta-catenin, 14-3-3 zeta, and glucose-6-phosphate dehydrogenase were found to colocalize with CK aggregates.
• E-cadherin exhibited altered localization in CK aggregate-containing cells.
• Cells with CK aggregates showed significantly increased size and altered morphology compared to controls.

Conclusions:

• Cytokeratin aggregates disrupt normal cellular architecture and protein localization.
• The observed changes in junction-associated and cytosolic proteins suggest impaired cellular function.
• These findings provide insights into the cellular consequences of protein aggregation in disease states.